Fig. 2: Transplanted enhancers maintain open chromatin structure and drive functional Shh expression in the limb at short range.
From: Range extender mediates long-distance enhancer activity
a, Allele-specific ZPA ATAC–seq profiles at the transplanted HS1516 (green box) and HS72 (orange box) enhancers and the corresponding WT ZRS locus (top). Bottom, ATAC–seq profiles at endogenous mHS1516 (left; chromosome 16: 95847911–95849564; mm10) and mHS72 (right; chromosome 8: 89454508–89455383; mm10) enhancers (green and orange striped boxes). b, LacZ-stained transgenic E11.5 mouse embryo carrying the HS72 limb enhancer upstream of the Shh promoter (light blue) and lacZ reporter gene (dark blue). The number of embryos with robust LacZ staining in limb buds over the total number of transgenic embryos screened is indicated. c, Schematic of the ShhHS72(+2kb) KI allele, in which the HS72 enhancer was inserted around 2 kb upstream of the Shh TSS. Skeletal preparation of a forelimb from ShhHS72(+2kb)/WT mouse is shown below on the left. The star symbols indicate extra digits (polydactyly). Hindlimb images are shown in Extended Data Fig. 4b. Quantitative PCR (qPCR) analysis of gene expression in anterior and posterior E11.5 embryonic hindlimb buds in ShhHS72(+2kb)/WT (n = 8 embryos) and WT (n = 6 embryos) mice. Expression is normalized to the average Shh expression in the WT ZPA. Statistical analysis was performed using two-sided Wilcoxon rank-sum tests with no adjustments for multiple comparisons; **P = 0.000311. Scale bars, 1 mm (b (left) and c) and 250 μm (b, right).
